Requirements for syrM and nodD Genes in the Nodulation of Medicago truncatula by Rhizobium meliloti 1021
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Medicago truncatula, a relative of alfalfa (Medicago sativa), has been proposed as a model plant system for study of its interaction with the symbiont Rhizobium meliloti. Differences in M. truncatula and alfalfa may result in distinct symbiosis behaviors for the two hosts when in association with wild-type or mutant bacterial strains. We have found that M. truncatula has a more stringent requirement for syrM and nodD3 than is seen in alfalfa. In particular, the lack of syrM is associated with arrested nodule development.Keywords:
Medicago truncatula
Medicago
Medicago sativa
Sinorhizobium meliloti
Medicago truncatula
Sinorhizobium meliloti
Medicago
Sinorhizobium
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The specific Sinorhizobium meliloti lipopolysaccharide (LPS) mutant Rm6963 (A. Lagares, G. Caetano Anolles, K. Niehaus, J. Lorenzen, H. D. Ljunggren, A. Puhler, and G. Favelukes, J. Bacteriol. 174:5941-5952, 1992) was shown to be mutated in a region corresponding to a cloned 5-kb SstI DNA fragment that was able to complement the lpsB and lpsC mutants of S. meliloti described by Clover et al. (R. H. Clover, J. Kieber, and E. R. Signer, J. Bacteriol. 171:3961-3967, 1989). Sodium dodecyl sulfate polyacryla-mide electrophoresis revealed that the LPS-I and LPS-II fractions of the LPS mutant Rm6963 were shifted to lower molecular weights. While the majority of the Medicago spp. tested established an effective symbiosis with both the S. meliloti wild-type Rm2011 and the LPS mutant Rm6963, the latter induced ineffective nodules on M. truncatula. A light- and electron-microscopic analysis of the ineffective M. truncatula root nodules revealed that the bacteria were released from the infection threads but failed to colonize the plant cells effectively. The plant cytoplasm was filled with numerous vesicles, probably the result of a disturbed bacteroid development. Sections of ineffective M. truncatula root nodules induced by the LPS mutant Rm6963 showed brown, necrotic cells within the central nodule tissue that autofluoresced when viewed under UV light. These observations are best explained by a plant defense response. Evidently, the rhizobial LPS plays a role in plant-microbe signaling during the formation of M. truncatula nodules.
Medicago truncatula
Sinorhizobium meliloti
Medicago
Sinorhizobium
Medicago sativa
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Host specificity in the root-nodule symbiosis between legumes and rhizobia is crucial for the establishment of a successful interaction and ammonia provision to the plant. The specificity is mediated by plant-bacterial signal exchange during early stages of interaction. We observed that a Sinorhizobium meliloti mutant ∆relA, which is deficient in initiating the bacterial stringent response, fails to nodulate Medicago sativa (alfalfa) but successfully infects Medicago truncatula. We used biochemical, histological, transcriptomic, and imaging approaches to compare the behavior of the S. meliloti ∆relA mutant and wild type (WT) on the two plant hosts. ∆relA performed almost WT-like on M. truncatula, except for reduced nitrogen-fixation capacity and a disorganized positioning of bacteroids within nodule cells. In contrast, ∆relA showed impaired root colonization on alfalfa and failed to infect nodule primordia. Global transcriptome analyses of ∆relA cells treated with the alfalfa flavonoid luteolin and of mature nodules induced by the mutant on M. truncatula revealed normal nod gene expression but overexpression of exopolysaccharide biosynthesis genes and a slight suppression of plant defense-like reactions. Many RelA-dependent transcripts overlap with the hypo-osmolarity-related FeuP regulon or are characteristic of stress responses. Based on our findings, we suggest that RelA is not essential until the late stages of symbiosis with M. truncatula, in which it may be involved in processes that optimize nitrogen fixation.
Sinorhizobium meliloti
Sinorhizobium
Medicago
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Nitrogen-fixing root nodules represent strong carbon sinks. Sugar partitioning was studied in three different symbiotic systems, namely a legume, Medicago truncatula, and two actinorhizal plants, Casuarina glauca and Datisca glomerata. The expression levels of sucrose synthase (SuSy) and genes encoding sugar transporters were compared in roots and nodules. Legume and actinorhizal symbioses differ with respect to hexose transporter (HT) expression: induction of HT genes was found in nodules of actinorhizal plants Datisca and Casuarina, but HT expression levels in Medicago nodules were strongly reduced compared to roots. Expression levels of sucrose transporter (ST) genes in Medicago nodules were strongly reduced compared to roots.Enzyme activities of three invertase isoforms (vacuolar, apoplastic and cytosolic) and SuSy was determined for roots and nodules. The sum of apoplastic, cytosolic, and vacuolar invertase activities is strongly reduced in nodules compared to roots of all three symbiotic systems studied. The major sucrose-cleaving activity in Medicago and Datisca roots and nodules is represented by SuSy. Extracts from Datisca and Casuarina contain a single immunoreactive SuSy protein band of about 92 kDa. Medicago contains an additional, somewhat larger SuSy isoform in roots, but not in nodules. SuSy activities were similar in soluble protein fractions from both roots and nodules of Medicago, while in Datisca nodules they were lower than in roots.The profiles of soluble sugars analysed in roots and nodules. Two unknown sugars were isolated from Datisca and identified using biochemical methods, NMR and mass spectroscopy. They represented rutinose (α-L-rhamnopyranosyl-(1→6)-D-glucose) and methylrutinose (α-L-rhamnopyranosyl-(1→6)-1-O-methyl-β-D-glucose). Both were present in high amounts in roots, nodules and leaves of Datisca. Sucrose is the major sugar of roots and nodules from Medicago and Casuarina, while the major sugar of Datisca roots and nodules is rutinose.Full-size hexose transporter cDNAs were isolated from Medicago (MtHT) and Datisca (DgHT) nodules. Based on their amino acid sequence, the encoded proteins belonged to different classes of sugar transporters. The functional characterization of DgHT in a heterologous system (yeast) demonstrated that DgHT is a high affinity, energy-dependent monosaccharide transporter with broad substrate specificity, probably an H+-symporter.The implications of these data on sugar partitioning mechanisms are discussed.
Medicago truncatula
Frankia
Actinorhizal plant
Medicago
Leghemoglobin
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Medicago truncatula
Sinorhizobium meliloti
Sinorhizobium
Medicago
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新兴的证据证明了 ROP GTPases 在共生起重要作用，但是他们在共生的规定上的分子的机制大部分是未知的。在这研究，我们证明 MtROP8 涉及在 Medicago truncatula 和 Sinorhizobium meliloti 之间的共生相互作用。表示分析证明 MtROP8 在早感染的根是下面调整的，但是在与根相比的小瘤显著地起来调整。 RNA 干扰( RNAi )的 Phenotypic 分析调停了 MtROP8 的 silencing 揭示了那 MtROP8 击倒表示导致了根头发的各种各样的发展缺点包括分叉的头发，短球状的根头发，和有明显的肿的底的甚至根头发，它被分发的修正和反应的氧种类( ROS )的层次引起。而且，感染事件在响应 S 怀有 MtROP8 RNAi 构造的转基因的根被增加。meliloti 接种，有提高的生节的伴随物。这些结果显示 MtROP8 由调整 ROS 生产和分发参予根头发开发和共生相互作用的建立。
Medicago truncatula
Sinorhizobium meliloti
Sinorhizobium
Medicago
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MtN6 belongs to a series of cDNA clones representing Medicago truncatula genes transcriptionally activated during nodulation by Sinorhizobium meliloti (P. Gamas, F. de Carvalho Niebel, N. Lescure, and J. V. Cullimore, Mol. Plant-Microbe Interact. 9:233–242, 1996). We show here by in situ hybridization that MtN6 transcripts specifically accumulate first at very localized regions in the outer root cell layers, corresponding to outer cortical cells containing preinfection threads. At later stages, MtN6 expression is observed ahead of growing infection threads, including in the infection zone of mature root nodules. Interestingly, regulation of MtN6 is clearly distinct from that of other early nodulins expressed in the same region of the nodule, in terms of response to bacterial symbiotic mutants and to purified Nod factors. We thus suggest that MtN6 represents the first specific marker of a pathway involved in preparation to infection, which is at least partly controlled by Nod factors. Finally, we discuss the intriguing sequence homology shown by MtN6 to a protein from Emericella (Aspergillus) nidulans, FluG, that plays a key role in controlling the organogenesis of conidiophores (B. N. Lee and T. H. Adams, Genes Dev. 8:641–651, 1994).
Medicago truncatula
Sinorhizobium meliloti
Sinorhizobium
Medicago
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The acidic polysaccharide succinoglycan produced by the nitrogen-fixing rhizobial symbiont Sinorhizobium meliloti 1021 is required for this bacterium to invade the host plant Medicago truncatula and to efficiently invade the host plant M. sativa (alfalfa). The β-glucanase enzyme encoded by exoK has previously been demonstrated to cleave succinoglycan and participate in producing the low molecular weight form of this polysaccharide. Here, we show that exoK is required for efficient S. meliloti invasion of both M. truncatula and alfalfa. Deletion mutants of exoK have a substantial reduction in symbiotic productivity on both of these plant hosts. Insertion mutants of exoK have an even less productive symbiosis than the deletion mutants with the host M. truncatula that is caused by a secondary effect of the insertion itself, and may be due to a polar effect on the expression of the downstream exoLAMON genes.
Medicago truncatula
Sinorhizobium meliloti
Medicago
Medicago sativa
Sinorhizobium
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Leguminous plants are able to form a root nodule symbiosis with nitrogen-fixing soil bacteria called rhizobia. This symbiotic association shows a high level of specificity. Beyond the specificity for the legume family, individual legume species/genotypes can only interact with certain restricted group of bacterial species or strains. Specificity in this system is regulated by complex signal exchange between the two symbiotic partners and thus multiple genetic mechanisms could be involved in the recognition process. Knowledge of the molecular mechanisms controlling symbiotic specificity could enable genetic improvement of legume nitrogen fixation, and may also reveal the possible mechanisms that restrict root nodule symbiosis in non-legumes. We screened a core collection of Medicago truncatula genotypes with several strains of Sinorhizobium meliloti and identified a naturally occurring dominant gene that restricts nodulation by S. meliloti Rm41. We named this gene as Mt-NS1 (for M. truncatula nodulation specificity 1). We have mapped the Mt-NS1 locus within a small genomic region on M. truncatula chromosome 8. The data reported here will facilitate positional cloning of the Mt-NS1 gene. Evolution of symbiosis specificity involves both rhizobial and host genes. From the bacterial side, specificity determinants include Nod factors, surface polysaccharides, and secreted proteins. However, we know relatively less from the host side. We recently demonstrated that a component of this specificity in soybeans is defined by plant NBS-LRR resistance (R) genes that recognize effector proteins delivered by the type III secretion system (T3SS) of the rhizobial symbionts. However, the lack of a T3SS in many sequenced S. meliloti strains raises the question of how the specificity is regulated in the Medicago-Sinorhizobium system beyond Nod-factor perception. Thus, cloning and characterization of Mt-NS1 will add a new dimension to our knowledge about the genetic control of nodulation specificity in the legume-rhizobial symbiosis.
Medicago truncatula
Sinorhizobium meliloti
Sinorhizobium
Medicago
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In this study, we investigated genetic elements of the type IV secretion system (T4SS) found in Sinorhizobium spp. and the role they play in symbiosis. Sinorhizobium meliloti and S. medicae each contain a putative T4SS similar to that used by Agrobacterium tumefaciens during pathogenesis. The Cre reporter assay for translocation system was used to validate potential effector proteins. Both S. meliloti and S. medicae contained the effector protein TfeA, which was translocated into the host plant. Sequence analysis revealed the presence of a nod box involved in transcriptional activation of symbiosis-related genes, upstream of the transcriptional regulator (virG) in the Sinorhizobium T4SS. Replicate quantitative reverse transcription-polymerase chain reaction analyses indicated that luteolin, released by roots and seeds of Medicago truncatula, upregulated transcription of tfeA and virG. Mutations in the T4SS apparatus or tfeA alone resulted in reduced numbers of nodules formed on M. truncatula genotypes. In addition, S. meliloti KH46c, which contains a deletion in the T4SS, was less competitive for nodule formation when coinoculated with an equal number of cells of the wild-type strain. To our knowledge, TfeA is the first T4SS effector protein identified in Sinorhizobium spp. Our results indicate that Sinorhizobium i) uses a T4SS during initiation of symbiosis with Medicago spp., and ii) alters Medicago cells in planta during symbiosis. This study also offers additional bioinformatic evidence that several different rhizobial species may use the T4SS in symbiosis with other legumes.
Medicago truncatula
Sinorhizobium meliloti
Sinorhizobium
Medicago
Medicago sativa
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